EP3527653A1 - Système d'exposition et procédé de gazage continu d'au moins une culture cellulaire - Google Patents

Système d'exposition et procédé de gazage continu d'au moins une culture cellulaire Download PDF

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Publication number
EP3527653A1
EP3527653A1 EP18157634.9A EP18157634A EP3527653A1 EP 3527653 A1 EP3527653 A1 EP 3527653A1 EP 18157634 A EP18157634 A EP 18157634A EP 3527653 A1 EP3527653 A1 EP 3527653A1
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EP
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Prior art keywords
gas
water
bar
standard
test gas
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EP18157634.9A
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German (de)
English (en)
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EP3527653B1 (fr
Inventor
Johanna Gostner
Johannes Zeisler
Reinhold Steinlechner
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Fritz Egger GmbH and Co OG
Medizinische Universitaet Innsbruck
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Fritz Egger GmbH and Co OG
Medizinische Universitaet Innsbruck
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Priority to EP18157634.9A priority Critical patent/EP3527653B1/fr
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M29/00—Means for introduction, extraction or recirculation of materials, e.g. pumps
    • C12M29/26—Conditioning fluids entering or exiting the reaction vessel
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/30—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration
    • C12M41/32—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration of substances in solution
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/30—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration
    • C12M41/34—Means for regulation, monitoring, measurement or control, e.g. flow regulation of concentration of gas
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M41/00—Means for regulation, monitoring, measurement or control, e.g. flow regulation
    • C12M41/40—Means for regulation, monitoring, measurement or control, e.g. flow regulation of pressure
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M45/00—Means for pre-treatment of biological substances
    • C12M45/03—Means for pre-treatment of biological substances by control of the humidity or content of liquids; Drying

Definitions

  • the invention relates to an exposure system for the continuous gassing of at least one cell culture with at least one test gas, with at least one sample receptacle for keeping the at least one cell culture ready during the gassing and with at least one conditioning device for conditioning the at least one test gas, wherein the conditioning device comprises at least one carrier gas supply, at least one Testkomponentzugabe and at least one Testgasbefeuchtung and wherein the at least one Testgasbefeuchtung at least one water tank and / or the test component addition at least one standard container for holding at least one at least one at least one test component having standards and wherein the standard container at least one pressure supply for applying the dosed standard in Standard container is associated with a predetermined pressure. Furthermore, the invention relates to a method for operating an exposure system for continuously gassing at least one cell culture in at least one sample intake with at least one test gas.
  • Exposure systems with which cell cultures can be exposed to a controlled gas atmosphere, are known to varying extents.
  • the exposure systems serve to study the effects of certain substances on the human organism.
  • the substances tested with appropriate exposure equipment are typically those that are considered to be at least potentially toxic or carcinogenic.
  • these are in particular substances that are absorbed via the respiratory tract, such as Volatile Organic Compounds (VOCs).
  • VOCs Volatile Organic Compounds
  • the cell cultures are treated with a test gas with a certain concentration of the test Component flows over, as Zellkuturen example lung epithelial cells or immune cells in question.
  • the pressurized standard container is taken from a predetermined mass flow, which is controlled by a mass flow controller and then evaporated.
  • the standard is preferably a liquid or solution which has a certain predetermined proportion of a test component.
  • the test component can be dissolved or absorbed in the standard.
  • a certain amount of test component can be provided for gassing the cell cultures.
  • the standard could in principle also be the test component in its purest form. However, the dosage is simpler and more accurate because of the small amount of test component required if the test component is only a small part of the standard.
  • the standard may, if necessary, also have several test components with which the cell cultures are to be fumigated at the same time. However, an exact, permanent adjustment of the test gas composition has also been found to be unsatisfactory.
  • the object of the present invention is to design and further develop the exposure system and the method of the type mentioned above and described in more detail above in such a way that reliable long-term measurements with exposure systems can be carried out.
  • the water tank is provided at least a pressure supply for applying the test gas to be metered water in the water tank with a predetermined pressure that the water tank a water capillary for passing the metered and with downstream of the predetermined pressure of the pressurized water and that the water capillary an evaporator for evaporating the water capillary passing water is connected downstream.
  • the invention has recognized that a more uniform and accurate dosing of the moisture and the test component or of the standard can take place if the mass flow controller is dispensed with in favor of a capillary.
  • Mass flow controllers are typically particularly suitable for permanently regulating a mass flow very precisely. In the case of the very low mass flows, however, this is only possible to a limited extent.
  • the carrier gas is passed through a wash bottle after passing through the mass flow controller.
  • the moisture absorption does not appear to be accurate and constant here despite a temperature of the wash bottle. This may be related to fluctuations in the mass flow of carrier gas, but should also be largely conditioned by temperature fluctuations in the wash bottle.
  • the invention enables a highly interference-free and accurate adjustment of the test component.
  • the standard is pressed through a liquid capillary on the standard container under a certain pressure as in the water tank and then completely evaporated in an evaporator.
  • a liquid capillary makes it possible to set the mass flow very accurately over the pressure of the standard in front of the liquid capillary.
  • test component addition and the test gas humidification can also be used independently of each other. Both the test component addition and the test gas humidification each lead to the advantages mentioned above. However, it is particularly preferred if test component addition and the test gas humidification are configured as described in order then to be able to combine the corresponding advantages.
  • the test component may in principle be very different substances. Nor is the invention limited to the use of a single test component. It is also possible to use several test components simultaneously for the formation of a test gas. Preferably, a standard is then used in which all test components are contained in proper proportion to each other to obtain a test gas having the desired test gas composition. However, different standards can also be kept in different standard containers. Then the dosage of each individual standard can be done as previously described. Each standard will then pressed through a separate liquid capillary and then evaporated in a separate evaporator. The individual vaporized standards can then be subsequently combined in a mixer or in several mixers. If necessary, a vapor capillary for buffering the pressure and the mass flow may be provided between the at least one mixer and at least one evaporator.
  • the at least one test component is particularly preferably a volatile organic compound (VOC), since knowledge about its effects on the human organism is of particular interest. This applies, for example, to formaldehyde (CH 2 O) and many other test components.
  • VOC volatile organic compound
  • epithelial cells are in monoculture or co-culture with other cell types.
  • Such co-culture may preferably be co-culture lung epithelial cells with immune cells to simulate the uptake of potentially hazardous substances via the respiratory tract.
  • the invention allows a preferred operation of a conditioning device for conditioning a test gas for gassing the at least one cell culture in the exposure system in which water to be metered from at least one pressurized water container is supplied with a predetermined pressure to a water capillary and the water is evaporated after leaving the water capillary ,
  • the standard to be dispensed from at least one pressurized standard container can be supplied with a predetermined pressure to a liquid capillary and the standard evaporated after leaving the liquid capillary.
  • the pressure supply of the water tank and / or the standard container has at least one compressed gas connection, in particular compressed air connection.
  • a constant pressure in the water tank and / or standard container can be easily and permanently maintained.
  • at least one pressure control is provided for regulating the pressure in the water container and / or in the standard container.
  • a reducing valve can be provided in the compressed gas line.
  • the pressure in the water tank and / or in the standard container between 1 bar and 4 bar, preferably between 1.1 bar and 3 bar, in particular between 1.2 bar and 2 bar, more particularly between 1.3 bar and 1.5 bar, is.
  • the evaporator for evaporating the water and / or the standard can be assigned at least one evaporator control for regulating the temperature of the evaporated water and / or the vaporized standard.
  • the evaporator control can alternatively or additionally also the Regulate the temperature of the evaporator itself, if this is expedient and the temperatures of evaporator and vaporized Waser or standard in practice differ significantly.
  • the temperature of the evaporated water and / or standards in the evaporator can preferably be adjusted to a specific value between 100 ° C. and 300 ° C., preferably between 120 ° C. and 200 ° C., in particular between 130 ° C. and 170 ° C. be managed.
  • the pressure of the evaporated water before the steam capillary and / or the vaporized standard before the vapor capillary to a certain value between 1 bar and 4 bar, preferably between 1.1 bar and 3 bar, in particular between 1.2 and 2 bar, in particular between 1.3 bar and 1.5 bar, set or regulated. This allows complete and even evaporation of the water and / or the standard. It also ensures that no unwanted condensation of moisture and / or standard occurs prior to loading the cell cultures with the test gas.
  • the mixer of the water, the evaporator of the standard, the Wasserdampfkapillare and / or the Dampfkapillare a mixer downstream be, which may be a static mixer in a particularly preferred embodiment.
  • the mixer can both serve to mix the evaporated water and the vaporized standard with each other and also be used in addition or simultaneously for mixing the evaporated water and / or the vaporized standard with the carrier gas.
  • the mixing of the individual streams to the test gas can thus be accomplished by a single mixer or by a plurality of mixers, in particular static mixers.
  • the carrier gas supply may have at least one gas connection, at least one air connection and / or carbon dioxide connection (CO2), in order to supply appropriate gas.
  • a mass flow controller mass flow controller
  • the at least one gas port and / or the at least one mass flow meter downstream of a distribution device for uniformly dividing the carrier gas for gassing of different cell cultures with different test gas concentrations. So not every single carrier gas flow must be set via separate mass flow meter. For this purpose, in particular offer critical nozzles, each having a very small opening.
  • an exposure system can be provided which comprises a plurality of cell boxes for gassing cell cultures with different test gas concentrations, and accordingly a plurality of test gas humidities and test component additions.
  • a carrier gas supply can then be provided with a distribution device which provides carrier gas streams for each of the cell boxes.
  • the at least one mass flow controller preferably regulates the mass flow of the carbon dioxide supplied via the gas connection to a constant, predetermined value between 5 Nl / h (standard liters per hour) and 100 Nl / h, preferably between 10 Nl / h and 50 Nl / h, in particular between 15 Nl / h and 30 Nl / h.
  • the at least one mass flow controller can control the mass flow of the supplied air to a constant, predetermined value between 100 Nl / h and 1200 Nl / h, preferably between 200 Nl / h and 800 Nl / h, in particular between 300 Nl / h and 500 Nl / h, regulate.
  • Nl thus represents a volume under standard conditions
  • the standard conditions (technical standard state) are given here by a standard pressure of 1013.25 mbar and a standard temperature of 293.15 K.
  • the at least one sample receptacle may be provided for keeping the at least one cell culture in a tempered exposure chamber during the fumigation. If the temperature control for controlling the temperature of the exposure chamber to a temperature between 34 ° C and 38 ° C, preferably between 35 ° C and 37 ° C, in particular of at least substantially 36 ° C, is provided, the temperature of the human lung, for example, can be simulated , In addition, drying out with high cell activity can be avoided.
  • the test gas can be supplied at a temperature above the dew point. There is then no risk of undesirable condensation.
  • the test gas can be passed into the exposure chamber with a very high relative humidity, so that the cell cultures do not dry out.
  • the relative humidity should in contact with the cell cultures quite basically in a preferred manner between 95% and 100%, but without it comes to condensation.
  • the upper limit would therefore be more preferably 99.5% relative humidity.
  • Favorable humidities are favorable for lung epithelial cells between 97% and 99%.
  • the target size can therefore be 98% relative humidity.
  • the at least one sample receptacle for keeping ready the at least one cell culture can be provided for better temperature control during the gassing in a cell box through which the test gas flows, which is located within the temperature-controlled exposure chamber.
  • the cell box can be hermetically separated from a temperature control area of the exposure chamber to preclude mixing between the test gas and the atmosphere of the exposure chamber.
  • at least one temperature sensor for detecting the temperature of the test gas in the cell box can be provided, so that the temperature of the test gas in the cell box can be adjusted very accurately on contact with the cell cultures or deviations from the desired state can be detected and detected.
  • a dew point temperature measurement may also be carried out in order to detect the dew point in the cell box and ultimately be able to regulate the dew point. This can be done for example via a variation of the pressure of the at least one water tank.
  • a control for adjusting the temperature of the test gas in the cell box between 35 ° C and 39 ° C, preferably between 36 ° C and 38 ° C, in particular 37 ° C ( ⁇ 0.5 ° C), be provided.
  • the nominal value of the dew-point temperature may be between 34 ° C. and 39 ° C., preferably between 36 ° C. and 37 ° C., in particular 36.5 ° C.
  • the at least one cell box has on the inlet side and / or outlet side a widening diffuser and / or a tapering diffuser.
  • the flow cross-section expands slowly upon entry into the cell box and tapers slowly as it exits the cell box without causing dead zones or particulate matter Turbulence would come.
  • the at least one cell box above the at least one sample receptacle may have at least one grid network and / or perforated plate overflowed by the test gas. There is then a tendency to constant concentration of test components in the test gas, so that also forms a driving concentration difference for the test concentration from there to the cell contours.
  • At least one temperature-controlled sampling line can be provided. This can then be connected to an analyzer, which determines the gas composition.
  • a condensation line for condensing vaporous portions of the gas may be provided in a condenser.
  • the pressure in at least one water tank and / or standard container preferably via at least one compressed gas connection and / or at least one reducing valve, regulated.
  • This allows an accurate and constant release of water and / or standard to produce a test gas that has exactly the desired composition and also retains it.
  • This is done very precisely and efficiently when the pressure in the water tank and / or in the standard container is constant at a value between 1 bar and 4 bar, preferably between 1.1 bar and 3 bar, in particular between 1.2 bar and 2 bar, more particularly between 1.3 bar and 1.5 bar, set, in particular, is regulated.
  • the temperature of the evaporated water and / or the vaporized standard in the evaporator to a temperature between 100 ° C and 300 ° C, preferably between 120 ° C and 200 ° C, in particular between 130 ° C and 170 ° C, adjusted, in particular, be controlled.
  • the pressure of the evaporated water before the steam capillary and / or the pressure of the vaporized standard before the vapor capillary to a pressure between 1 bar and 4 bar, preferably between 1.1 bar and 3 bar, in particular between 1.2 bar and 2 bar, more particularly between 1.3 bar and 1.5 bar, adjusted, in particular, be regulated.
  • the corresponding capillary has a buffer effect or equalizing effect with regard to the mass flow after the corresponding capillary.
  • At least one mixer in particular a static mixer, may be connected downstream of the evaporator of the water, the evaporator of the standard, the steam capillary and / or the vapor capillary.
  • a static mixer may be connected downstream of the evaporator of the water, the evaporator of the standard, the steam capillary and / or the vapor capillary.
  • the evaporated water, the vaporized standard and / or the carrier gas is mixed. All together then form the test gas, with only two of the mass flows mentioned mixed in the mixer or the mass flows in more than one mixer, if necessary, one after the other, can be mixed.
  • the mixing together in a single mixer is particularly simple in terms of apparatus and nevertheless, in particular when using a static mixer, results in a uniform test gas.
  • a predetermined mass flow of air and / or of carbon dioxide (CO 2) can be supplied, in particular mixed with one another.
  • This serves to provide a suitable carrier gas as part of the test gas.
  • the carbon dioxide with a certain mass flow to be between 5 Nl / h and 100 Nl / h, preferably between 10 Nl / h and 50 Nl / h, in particular between 15 Nl / h and 30 Nl / h, and / or the air with a certain mass flow between 100 Nl / h and 1200 Nl / h, preferably between 200 Nl / h and 800 Nl / h, in particular between 300 Nl / h and 500 Nl / h, is supplied.
  • the supply preferably follows a mixing in a mixer, which can be designed simply and effectively as a static mixer.
  • a mass flow controller is particularly suitable.
  • the at least one mass flow meter can be followed by a distributor for uniformly dividing the carrier gas for gassing of different cell cultures with different test gas concentrations, so that not every single carrier gas flow has to be set via separate mass flow meters.
  • a distributor for uniformly dividing the carrier gas for gassing of different cell cultures with different test gas concentrations, so that not every single carrier gas flow has to be set via separate mass flow meters.
  • critical nozzles each having a very small opening. The opening is the same for all critical nozzles, so that the flow through the critical nozzles can be divided into exactly equal proportions.
  • Meaningful measurement can be achieved without drying of the cell cultures, if the at least one sample intake during the gassing of the corresponding cell culture in one, preferably at a certain temperature between 34 ° C and 38 ° C, preferably between 35 ° C and 37 ° C, in particular is kept at least substantially 36 ° C, tempered exposure chamber.
  • the test gas can be heated to a specific temperature immediately before being guided to a cell box arranged in the exposure chamber between 36 ° C and 40 ° C, preferably between 37 ° C and 39 ° C, in particular of at least substantially 38 ° C, tempered, in particular cooled.
  • the test gas is not brought close to the target temperature until immediately before entering the arranged in the exposure chamber cell box, so that premature condensation can be reliably prevented.
  • FIG. 1 An exposure analysis 1 with an exposure chamber 2 is shown, in which cell cultures can be gassed with a test gas of a specific, predetermined test gas composition.
  • the composition of the test gas is adjusted and regulated in the conditioning device 3, so that the test gas composition is not subject to appreciable fluctuations even over very long periods of many days.
  • the conditioning device 3 comprises a test component addition 4, via which the desired amount of a test component is added to a test gas.
  • the test component is in the form of a so-called standard 5, for example in pure form or in a certain concentration in a solution or absorbed in an absorbent before.
  • the concentration of the test component is known very accurately and set very precisely to a certain value.
  • the standard 5 is in liquid form, which greatly simplifies its handling.
  • the standard 5 is kept in a standard container 6, which is connected via a gas connection 7 to a gas supply 8.
  • the gas supply 8 can are formed for the sake of simplicity by a compressed gas tank.
  • the gas of the gas supply 8 may be compressed air for the sake of simplicity.
  • the pressure with which the standard container 6 is acted upon via the gas connection 7 can be set, in particular regulated.
  • the pressure is present between 1.3 bar and 1.5 bar.
  • the temperature of the standard container 6 also corresponds, if necessary, room temperature.
  • the standard can now be passed with the pressure of the standard container 6 in a liquid capillary 11, which ensures that a directly connected to the pressure of the standard container 6 and therefore adjusted very accurately mass flow of standard 5 in one of the liquid capillary 11 downstream Evaporator 12 is passed.
  • a temperature of about 150 ° C and a pressure of 1.3 bar to 1.5 bar, so that the standard 5 is completely evaporated in the evaporator 12.
  • Downstream of the evaporator 12 is a vapor capillary 13, which ensures buffering of the pressure and the mass flow to the evaporator 12.
  • the liquid capillary 11 has a smaller diameter and a greater length than the vapor capillary 13.
  • the mass flow of standard 5 may be, for example, between 1 g / h and 50 g / h, but further preference is given to mass flows of between 2 g / h and 25 g / h, in particular between 3 g / h and 10 g / h. In the present case, the mass flow of the standard is about 5 g / h.
  • the liquid capillary 11 and the vapor capillary 13 may, for example, have a diameter of between 0.1 mm and 0.8 mm and a length of up to 5 cm.
  • the diameter of the liquid capillary 11 is preferably about 0.1 mm, while the diameter of the vapor capillary 13 is preferably about 0.8 mm.
  • the separate carrier gas supply 14 in the present case has two gas connections 15, 16, it being possible in principle for the number of gas connections to vary.
  • a gas connection may be sufficient, for example if the gas connection already provides the carrier gas in the desired composition or a pure gas is to be used as the carrier gas.
  • a carrier gas comprising air and carbon dioxide is provided via the carrier gas feed 14. Therefore, the gas ports are also formed as a compressed air port and as a carbon dioxide port. Again, the air and the carbon dioxide for the sake of simplicity again about compressed gas tank 8,17 containing on the one hand air and on the other hand carbon dioxide provided.
  • the air and the carbon dioxide are cleaned after being drawn off via the respective gas connections 15, 16 by two high-temperature sterile filters 18 in each case.
  • the high-temperature sterile filter 18 are heated to 180 ° C to 200 ° C to keep the high-temperature sterile filter 18 sterile, the gas is much less heated due to the very short residence time in the high-temperature sterile filter 18.
  • the air and the carbon dioxide are then each fed to a mass flow controller 19, via which the respective mass flow of the air and of the carbon dioxide is regulated.
  • the mass flow is about 20 Nl / h (standard liters per hour), while the mass flow of air is about 380 Nl / h.
  • the gas streams are combined after the mass flow controllers 19,20 in a manifold 21.
  • the mass flow controllers 19, 20 are followed by a distributor device 22 for uniformly dividing the carrier gas for gassing of different cell cultures with different test gas concentrations, which, however, is by no means compulsory.
  • the carrier gas of the adjusted concentrations can be divided evenly over the distributor 22 without the need for further mass flow meters.
  • the individual carrier gas streams can then be mixed with different concentrations of the test component in order to gas different cell cultures, if necessary of the same type, in parallel with different test gas concentrations. Then, not every single carrier gas flow has to be set via separate mass flow meters.
  • the distributor 22 has a number of parallel connected critical nozzles, which corresponds to the number of individual carrier gas streams.
  • the critical Nozzles each have a very small opening, the openings being the same size for all critical nozzles, so that the flow can easily be divided into exactly equal proportions via the critical nozzles.
  • an exposure system can be provided which comprises a plurality of cell boxes 2 for gassing cell cultures with different test gas concentrations and accordingly a plurality of test component additions 4 and test gas humidifications 23.
  • only one carrier gas feed 14 can then be provided with a distribution device 22, which provides carrier gas streams for each of the cell boxes 2.
  • a test gas humidification 23 sets the desired moisture content of the test gas.
  • the water for humidifying the carrier gas is provided in the present case via a reservoir 24, from which then the water tank 25 feeds. But it could also be provided instead of the reservoir 25 only a water connection.
  • the water tank 25 is pressurized via a compressed gas connection 26, which in the present case is a compressed air connection, which can be adjusted and / or regulated via a reducing valve 27.
  • the pressure of the water tank 25 is in the present case between 1.3 bar and 1.5 bar and presses the water through a shut-off 28 through a water capillary 29, which generates such a flow resistance that the pressure in the water tank 25 very precisely the desired mass flow of water Humidifying the carrier gas can be adjusted.
  • the corresponding mass flow of water may for example be between 4 g / h and 80 g / h, preferably between 8 g / h and 40 g / h, in particular between 12 g / h and 25 g / h.
  • the mass flow of water is about 16 g / h, which passes after the water capillary 29 in an evaporator 30 in which a temperature before about 150 ° C and a pressure between 1.3 bar and 1.5 bar prevails.
  • the water capillary 29 has a smaller diameter and a greater length than the water vapor capillary 31.
  • the water capillary 29 and the steam capillary 31 may for example have a diameter between 0.1 mm and 0.8 mm and a length up to 5 cm.
  • the diameter of the water capillary 29 is preferably about 0.1 mm, while the diameter of the water vapor capillary 31 is preferably about 0.8 mm.
  • the vaporized standard 5, the carrier gas and the water vapor are fed separately to a mixer 32 which is a static mixer and in which a homogeneous test gas having a homogeneous test gas composition is produced.
  • the exposure unit 1 is not limited to the use of a single mixer 32 for this purpose.
  • a test gas temperature control for adjusting the temperature of the test gas.
  • a tempering device 33 for example in the form of an airbag, is provided which tempers the test gas to a temperature just above the dewpoint temperature. In the present case, the test gas is tempered immediately before the exposure chamber 2 to about 38 ° C.
  • the test gas is then introduced into a cell box 34, which is located in the temperature-controlled exposure chamber 2.
  • the exposure chamber 2 is tempered in the present case to a temperature of about 36 ° C.
  • a heating device 35 with a fan for equalizing the temperature in the exposure chamber 2 is provided in the illustrated and so far preferred exposure chamber 2.
  • the test gas in the exposure chamber 2 continues to cool, in such a way that the temperature of the test gas in the region of the cell cultures is about 37 ° C and thus has a relative humidity of about 98%.
  • the temperature of the test gas in the region of the cell cultures is about 37 ° C and thus has a relative humidity of about 98%.
  • two temperature measuring points are provided in the cell box 34, one in the test gas before reaching the cell cultures and one in the test gas after the overflow of the cell cultures. Roof that Overflowing the cell cultures, a dew point measurement can be carried out additionally or alternatively to the temperature measurement.
  • the nominal value of the dew point temperature at this point is 36.5 ° C.
  • a regulation ensures that the pressure in the water tank 25 is raised, as long as the dew point temperature falls below the setpoint temperature, and that the pressure in the water tank 25 is lowered when the dew point temperature exceeds the setpoint temperature.
  • test gas leaves the cell box 34 and the exposure chamber 2 and enters a heated chamber 36 whose temperature is preferably well above the dew point of the test gas to avoid premature condensation.
  • used test gas can be delivered to an analyzer 37 to determine the composition of the test gas used.
  • Test gas not used for analysis purposes is fed to a condenser 38, in which the condensable fractions of the test gas used, in particular the moisture and the test component, are condensed and thus deposited in a collecting container 39.
  • a heater 46 may optionally be provided with fan to equalize the temperature in the heated chamber 36.
  • the illustrated conditioning device 3 may be provided at least in part several times and connected to a respective separate exposure chamber 2.
  • the conditioning of different test gases can be done very accurately and at least substantially independently of the conditioning of other test gases.
  • Particularly preferred is the use of 4, 6 or 8 different exposure chambers 2, which can be operated with test gas of different concentration of test components, in a single exposure system.
  • all conditioning devices can, if required, be connected to a common gas connection, in particular compressed air connection and / or carbon dioxide connection.
  • a common storage tank for water can be used to fill or supply the water tank with water from this.
  • the carrier gas supply is simply provided. This applies in any case as long as the carrier gas produced as required from a plurality of gases by mixing in the carrier gas supply can be divided exactly onto the individual exposure chambers or onto the mixers for mixing the carrier gas with steam and vaporized standard.
  • test gas with a different, predetermined test gas composition is generated in each case. This can be achieved with appropriate control technology and equipment required, if necessary, by means of a single test gas supply. On the other hand, a single test component addition and a single test gas humidification would rather not be enough to simultaneously generate test gases with different test gas compositions.
  • FIG. 2A-B An aerodynamically optimized cell box 34 for receiving the cell cultures 40 is in the Fig. 2A-B shown.
  • the cell box 34 has a plurality of successively and juxtaposed sample receptacles 41. Basically, a single sample holder 41 would be sufficient here. However, more reproducible results are obtained using multiple sample receptacles 41 in a cell box 34.
  • the sample holders 41 are open at the top, so that the sample holders 41 and thus the cell cultures 40 overflowing test gas can come into contact with the cell cultures 40 in the sample holders 41 without mechanically damaging the cell carcasses 40 or putting them under stress.
  • a diffuser 42 is provided with a continuously expanding flow cross-section.
  • the diffuser 42 also meshes 43 and / or perforated plates are also incorporated, which are extend transversely to the entire flow cross section and thus force the entire test gas to flow through.
  • the thus uniformized flow of the test gas now flows through another grid 44 and / or perforated plate, which is arranged above the sample holders 41.
  • a defined and moderate flow of test gas can be conducted to the sample receptacles 41 and re-derived after contact with the cell cultures 40.
  • the atmosphere above the cell cultures 40 remains as constant as possible and also corresponds as possible to the test gas composition above the grid 44 and / or perforated plate.
  • the used test gas then flows out of the cell box 34 via a shorter diffuser 45 at the exit of the cell box 34.

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Cited By (1)

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Publication number Priority date Publication date Assignee Title
DE102019128558A1 (de) * 2019-10-22 2021-04-22 Copps Gmbh Druckgasaufbereitungsanlage

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